This proposal focuses on elucidation of the processes regulating human natural killer (NK) cell development. Our specific aims are: 1) To define the microenvironment(s) and characterize the regulation of NK cell differentiation in human secondary lymphoid tissues (SLT); and 2) To define the order, regulation, and phenotypic impact of major histocompatibility complex molecule binding receptor (MBR) acquisition during human NK cell maturation. NK cells are innate lymphoid cells (ILC) that have direct cytotoxic function against cancer cells. NK cells play a vital role in immune surveillance against malignant transformation by complementing T cell immunity, and they can mediate an important graft-versus-leukemia effect in the setting of allogeneic hematopoietic stem cell transplantation for acute myeloid leukemia (AML). Our overall goal in this proposal is to gain a comprehensive understanding of the cellular and molecular components that regulate human NK cell development and function in order to best understand how they work and can be enhanced in the face of malignancy. In particular, in Aim 1 we will define the cellular microenvironment(s) within SLT in which human NK cell developmental intermediates (NKDI) reside and differentiate. We previously identified and characterized the full spectrum of NKDI in these tissues. Over the past five years, additional and closely-related ILC subsets have been discovered and characterized, and we have preliminary data to indicate that all ILCs derive from a common ILC progenitor population that is naturally restricted to SLT. We hypothesize that the cellular milieu within SLT influences NK cell differentiation from the common ILC progenitor, and we propose a series of experiments to test our hypothesis and also to elucidate the mechanism(s) by which SLT-derived ?helper? cell populations influence this pathway. The goal of these studies is to identify new therapeutic targets to boost immune function in the post-transplant setting for AML. In Aim 2 we have preliminary data that suggest that NK cell functional maturation occurs in a stepwise fashion associated with the orderly and coordinated acquisition of the activation receptor, NKp80, and two types of MBRs: CD94/NKG2A and killer immunoglobulin-like receptors (KIR). What is not yet known and will be tested in our proposed experiments is how this orderly acquisition of receptors is regulated and in turn how signaling through the receptors themselves influences NK cell functional maturation. The clinical importance of these studies lies in the fact that each of these surface NK cell receptors has the potential to be targeted for immune checkpoint regulation in order to enhance NK cell-mediated immune therapy against cancer.